This invention relates to a biocompatible, implantable material which is absorbed naturally in vivo with minimal immunological reaction, and to the methods for its production. More particularly, this invention relates to a superior collagenous bone matrix useful as a xenogenic implant for use as an osteogenic device, as a bone particle coating for implantable prostheses, as a delivery vehicle for the in vivo sustained release of protein, and as a scaffold for anchorage-dependent cells.
A biocompatible, implantable material that can be resorbed in vivo could be used to promote conductive bone growth, induce osteogenesis when combined with an osteoinductive protein, provide a substratum for in vivo or in vitro growth of anchorage-dependent cells, or serve as a carrier for the sustained release of, for example, a therapeutic drug or antibiotic. Such a material must be biocompatible, that is, not induce an immunogenic/inflammatory response in vivo. Its physical structure must allow cell infiltration, and it must have an in vivo resorption time appropriate for its function.
The potential utility of an osteogenic device capable of inducing endochondral bone formation in vivo has been recognized widely. It is contemplated that the availability of such a device would revolutionize orthopedic medicine, certain types of plastic surgery, and various periodontal and craniofacial reconstructive procedures.
The developmental cascade of bone differentiation in mammalian bone tissue is well documented in the art (Reddi, 1981, Collagen Rel. Res. 1:209-226. Though the precise mechanisms underlying the phenotypic transformations are unclear, it has been shown that the natural endochondral bone differentiation activity of bone matrix can be dissociatively extracted and reconstituted with inactive residual collagenous matrix to restore full bone inducing activity (Sampath et al., 78 Proc. Natl. Acad. Sci. USA 7599-7603 (1981). Recently, the protein factors hereafter referred to as osteogenic protein (OP) responsible for inducing osteogenesis have been purified, expressed in recombinant host cells, and shown to be truly osteoinductive when appropriately sorbed onto a matrix. (U.S. Patent Application No. 179,406).
Studies have shown that while osteoinductive proteins are useful cross species, the demineralized bone matrix heretofore required for inducing endochondral bone formation is species specific (Sampath and Reddi (1983) PNAS 80:6591-6594). Implants of demineralized, extracted xenogenic bone matrix and OP invariably have resulted in a strong inflammatory response that has inhibited osteogenesis, presumably due to immunogenic protein components in the bone matrix. Hence, successful osteoinduction to date has required the use of allogenic bone matrix. This restriction on osteogenic devices is a serious limitation with respect to human clinical use, as human bone is neither readily available nor cost effective.
EPO 309,241 (published 3/29/89, filed 9/22/88, priority 9/25/87) discloses a device for inducing endochondral bone formation comprising an osteogenic matrix extract, and a matrix carrier comprising 60-90% mineral component and 2-40% collagen.
U.S. Pat. No. 4,563,350, published Jan. 7, 1986, discloses a xenogenic osteogenic device comprising a bone-inducing extract that had been Purified by gel filtration and ion exchange chromatography, and a collagenous matrix composed of approximately 90% trypsinized bovine bone matrix and 10% bovine dermal collagen. Endochondral bone formation is said to require the presence of at least 10-15% dermal collagen in the disclosed matrix.
Deatherage et al., (1987) Collagen Rel. Res. 7:2225-2231, purport to disclose an apparently xenogenic implantable device comprising a bovine bone matrix extract that has been minimally purified by a one-step ion exchange column and highly purified human Type-I placental collagen.
The current state of the art of materials used in surgical procedures requiring conductive bone repair, such as the recontouring or filling in of osseous defects, is disclosed by Deatherage (1988) J. Oral Maxillofac. Surg. 17:395-359. All of the known implant materials described (hydroxyapatite, freeze-dried bone, or autogenous bone grafts) have little or no osteoinductive properties. Clearly, the ability to induce osteogenesis is preferred over bone conduction for most procedures. Even when bone conduction is the indicated procedure for bone repair, a matrix consisting of non-immunogenic, extracted, xenogenic bone collagen heretofore has not been developed.
U.S. Pat. No. 4,795,467 discloses a bone repair composition comprising calcium phosphate minerals (preferable particle size of 100-2,000 .mu.) and atelopeptide, reconstituted, crosslinked fibrillocollagen. It purports to be a non-antigenic, biocompatible, composition capable of filling bony defects and promoting bone growth xenogenically.
U.S. Pat. No. 4,789,663 discloses a method of effecting conductive bone repair comprising exposing the defect to fresh bone, and using xenogenic collagen from bone and/or skin, wherein the collagen is enzymatically treated to remove telopeptides, and is artificially crosslinked.
The need to provide a "biological anchor" for implanted prostheses, particularly metallic implants, is well documented in the art. The state of the art of prosthetic implants, disclosed by Specter (1987) J. Arthroplasty 2:163-177, generally utilizes porous coated devices, as these coats have been shown to promote cellular ingrowth significantly.
EPO 169,001 (published 1/22/86, priority 07/17/84) claims a collagen-coated prosthesis wherein the coat comprises a purified, sterile, non-immunogenic xenogenic collagen preparation from bone or skin. Crosslinking is generally induced to reduce immunogenicity, or occurs as a result of sterilization procedures.
U.S. Pat. No. 4,812,120 discloses a prosthetic device comprising a metal core over which are applied successive polymers layers. The outer layer comprises a biopolymer having protruding collagen fibrils. The protruding fibrils are subject to damage upon implantation of the device.
Efficient in vitro growth of mammalian cells is often limited by the materials used as the substration, substratum, or "scaffold" for anchorage-dependent cells. An effective matrix must be physiologically acceptable to the anchorage dependent cells, and it must also provide a large available surface area to which the cells can attach.
GB Patent No. 2,178,447, published 02/11/87, claims a fibrous or porous foam matrix comprising open or closed form fibers, with a pore size on the order of 10-100 .mu. (matrix height is 50-500 .mu.). The fiber network is generated as a sheet which must then be modified if different scaffold shapes are desired.
Strand et al. (Biotechnology and Bioengineering, V. 26, 503, 1984) disclose microcarrier beads for use as a matrix for anchorage dependent cells in a matrix perfusion cell culture. Bead materials tested were DEAE or polyacrylamide. Surface area available was 250-300 cm.sup.2/ g and required a cell innoculaton of 10.sup.6 cells/ml.
U.S. Pat. No. 4,725,671 claims collagen fiber membranes suitable for cell culture, comprising soluble atelopeptide collagen fibers that are dried and preferably cross-linked.
The art has sought biocompatible sustained release vehicles with known, reliable "release" rates. Effective carriers must be biocompatible, water-insoluble, capable of trapping or otherwise holding the therapeutic agent of interest, and have a resorption time in vivo that mimics the desired release rate of the agent. Collagen and gelatin are attractive carriers for clinical use, primarily because of their biocompatible and biodegradable properties. (See, for example, EPO 170,979, published 02/12/86, priority 8/07/84; and EPO 069,260, published 01/12/83, priority 6/25/81.) However, these biopolymers have undesirable crosslinking properties that can make efficient synthesis of appropriate carrier matrices difficult. (EPO 230,647 published 08/05/87, priority 12/27/85.)
It is an object of this invention to provide a biocompatible, in vivo biodegradable bone matrix, implantable in a mammalian host with no significant inhibitory immunogenic response. Another object is to provide a biocompatible, in vivo biodegradable matrix capable of combining with an osteoinductive protein to produce endochondral bone formation in mammals, including humans. Still other objects are to provide a superior material for coating implantable prothetic devices, to increase the cellular ingrowth into such devices, to provide a biocompatible, in vivo biodegradable matrix for use as a carrier of sustained-release pharmaceutical compositions, wherein the resorption rate of the matrix can be adjusted to match that of the pharmaceutical agent, and to provide a biocompatible, in vivo biodegradable matrix capable of acting as a scaffold or substratum for anchorage-dependent cells, wherein the surface area available for cell attachment can be adjusted. Yet another object of the invention is to provide a method for the production of such matrix material.
These and other objects and features of the invention will be apparent from the description, drawings, and claims that follow.